Graft outcome is determined by the response and fate of alloreactive T cells in the host. When naive T cells encounter antigen, they face a series of "decisions", including whether to proliferate and develop effector function or to remain quiescent, and whether or not to progress from an effector to a memory cell (i.e., survive) or to undergo apoptosis. The ability to induce transplantation tolerance depends upon understanding and controlling these events. The overall goal of this project is to use in vitro and in vivo assays to analyze single cell T cell behavior using wild-type and alloreactive TCR transgenic mice, and transplantation and related relevant models. Our theoretical framework is that two of the most important parameters, which regulate T cell outcome following antigen encounter are the availability of costimulatory signals and whether or not the cell divides. Data from our laboratory indicate that these early events in the alloimmnune response of a T cell may irrevocably determine its destiny, and thus the fate of an allograft. Thus, the overall theme of the project is to understand how proliferation and priming of T cells regulate susceptibility to the development of effector and regulatory cells and the induction and maintenance of translantation tolerance. Along this theme, we have three aims. Aim #I will pursue exciting new preliminary data from our lab, suggesting the hypothesis that T cells undergoing homeostatic proliferation are resistant to transplantation tolerance induction. We will test this hypothesis in detail and study mechanisms responsible for this finding. Aim #2 will study the roles of T cell priming in the induction of effector and regulatory cells. Here, the use of a new CD4 alloreactive TCR transgenic mouse model will allow us to ask specific questions about priming and regulation, and identify and monitor the fate and function of individual all ore active T cells under very defined conditions. Aim #3 will focus on a population of T cells which we have recently identified, i.e., those which activated in response to antigen, but fail to proliferate, even in the presence of exogenous IL-2. We will study the conditions, which promote the generation of these cells, their responses to other T cell growth factors, and their potential function as regulatory cells. In selected studies, the availability of the microarray/genomics core will allow us to determine the genetic profile of alloreactive cells with well-defined, yet distinct, functional behaviors. Together, these investigations should provide new insights into how alloimmune T cell responses are initiated and controlled, and lead to the creation of novel tools and strategies to induce, maintain, and monitor tolerance.